The invention concerns a method for intracellular manipulation of at least one deposited biological cell situated in a cultural medium adhering to a support area, wherein an opening is created in the membrane of the cell, and the cell interior is manipulated through this opening. The invention furthermore relates to an apparatus for manipulation of the cell interior of at least one biological cell having a cell membrane situated in a culture medium. The apparatus has an object carrier, which has at least one support area on which the cell is adherently depositable, a poration tool for opening the cell membrane, and at least one entry channel situated in the area of the poration tool for manipulation of the cell interior.
An apparatus of the type mentioned at the beginning is already known from Alberts, B. et al., Molecular Biology of the Cell, Third Printing, VCH Verlag (1995), page 212 ff, which has a hollow needle connected with a suction device and made of an electrical insulating material having an interior cavity which has an opening on the free end of the hollow needle. For opening the cell membrane, the opening situated at the free end of the hollow needle is set upon the exterior of the cell membrane in order then to create an underpressure in the interior cavity of the hollow needle by means of the suction device. Through this underpressure, a cell membrane piece situated in front of the opening of the hollow needle is torn out of the membrane formation. After introducing the opening into the cell membrane the hollow needle, engaging the edge of the cell membrane surrounding the opening, electrically insulates the cell fluid contained in the interior of the cell against the culture medium. The interior of the cell is manipulated through the opening introduced into the cell membrane. For example, the cell nucleus can be removed by sucking cell fluid into the hollow needle from the cell, and subsequently another cell nucleus can be installed through the opening into the cell interior.
The previously known process and the apparatus for implementing the process have the disadvantage that a micromanipulator is necessary for positioning the hollow needle on the cell. This results in a comparatively more complicated and expensive device. Moreover, the accessibility of the cells situated on the object carrier by the micromanipulator is greatly reduced. The process and the apparatus are, for this reason, only suited for an intracellular manipulation of individual, or at best for a simultaneous manipulation of a small number of cells situated on the object carrrier. At the same time, a costly, manual positioning of the hollow needle on the cell is necessary.
Arranging a cell floating in a culture medium between two large surface area electrodes, respectively spaced at a distance from the cell, and applying an electric voltage to these electrodes is also already known from Alberts, B. et al., Molecular Biology of the Cell, Third printing, VCH Verlag (1995), page 213. Moreover, the membrane of the cell is simultaneously opened at several places by electroporation so that the genetic material situated in the culture medium to be introduced into the cell can be diffused into the cell interior through the openings of the cell. In this connection, however, it is disadvantageous that such an introduction of genetic material is subject to statistical fluctuations and is influenced by various parameters such as the concentration of genetic material in the area of the cell, the size of the genetic material to be introduced, and the size of the openings created in the cell membrane. The previously known process consequently does allow any selective manipulation of the cell interior.